This small grant (R03) proposal will seek to establish the feasibility of using peripheral blood stem cell (PBSC) as surrogate delivery systems of gene expression to the central nervous -system and addresses Research Objective No. 5 outlined in this 1VIH PA, " Stem Cells, Tissue Repair and Cell Replacement in Aging." Neuroprotective strategies aimed at preventing the neuropathological and behavioral sequelae of neurodenerative disease and aging might be powerful therapeutically since they could be introduced to patients before the onslaught of a cascade of irreversible, terminal events. Cellular delivery of neurotrophic factors using genetically modified cells is a particularly attractive strategy that could provide sustained expression of a vital trophic factor that has been depleted. There are several compelling advantages for the rationale of using PBSC. PBSC, in contradistinction to bone marrow-derived stem cells, are easily mobilized and readily-harvested intravenously with existing technology and avoids the painful, more invasive procedure associated with bone marrow aspiration. Clinically applied gene transfer strategies that employ mature, non-transformed neural cells, e.g. astrocytes or neural stem cells in autologous transplantation studies suffer from the unavoidable requisite neurosurgical procedure to obtain the brain-derived cells. In contrast, PBSC can be acquired free of surgical intervention and represent a potentially renewable source of cells for repeated cycles of gene therapy. This study will evaluate the capacity of G-CSF-mobilized harvested CD34+-enriched, human peripheral blood stem cells to undergo controlled in vitro proliferation and expansion toward myeloid stem cell lines a (non-lymphoid stem cell lineage) and ultimately toward monocyte/microglial precursor cell lineage. The cultured PBSC- derived monocyte/microglial precursor cells will be transduced by recombinant retroviruses to express selection (Neo) and marker (EGFP) genes and a protype neurotrophic factor gene that encodes ciliary neurotrophic factor (CNTF).The transduced cells will then serve as surrogate delivery systems of gene expression to the brain and spinal cord. The optimal procedure for delivery o transduced human peripheral blood stem cells to the brains of nude rats that results in sustained viability and constitutive expression of the transgenes will be defined. CNS PBSC implantation via intravenous infusion, intrathecal injection, intraventricular injection and by intracerebral injection will be examined. The fate of the transplanted, - transduced stem cells in the nude rat brain will be characterized with regard to the pattern of cellular migration and in vivo expression of the transgenes. The successful demonstration of a peripheral blood stem cell delivery system to the CNS will have road application in the design of gene therapies to effect tissue repair and cell replacement in aging.